1. Field of the Invention
This invention relates to a method of reendothelializing the vascular passage of a patient, the lining of which has been substantially denuded of endothelial cells by virtue, for example, of procedures such as endarterectomy.
2. Related Applications
U.S. Ser. No. 742,086, filed June 6, 1985, by Williams and Jarrell, discloses a method for treating a synthetic or naturally occurring implant intended for implantation in a human patient comprising the steps of obtaining human microvascular endothelial cell rich tissue from that patient, separating microvascular endothelial cells from that tissue; and applying said microvascular endothelial cells onto said implant to provide at least about 50% or greater confluence of said cells on the surface of said implant to be treated.
U.S. Ser. No. 848,453, filed on Apr. 4, 1986 as a continuation- in-part of U.S. Ser. No. 742,086, discloses a method of treating an implant intended for implantation in a human patient, comprising the steps of providing a synthetic substrate material and treating that material with Type IV/V collagen to improve human endothelial cell adhesion, proliferation and morphology. In the preferred embodiment, the endothelial cells are derived from the endothelial cell rich tissue of the patient undergoing implantation.
U.S. Ser. No. 927,745, filed by Jarrell and Williams on Nov. 6, 1986, discloses a method of determining endothelial cell coverage on a prosthetic surface.
The disclosures of these three applications are hereby incorporated by reference.
3. Description of the Art
Atherosclerotic vascular disease remains the leading cause of death among Americans. As medical science has become more sophisticated, increasing use of invasive vascular procedures are being applied to obstructed vessels in the absence of effective preventive or therapeutic drug modalities. For example, the use of arterial endarterectomy as well as percutaneous balloon dilatation of vessels for pathologic stenosis have become routine hospital procedures. Although these and other procedures are often successful, a common complication after the procedures is the occurrence of vessel wall abnormalities. These abnormalities include recurrent stenosis due to atherosclerosis, smooth muscle proliferation, loss of vessel wall integrity as a result of fibrosis and thrombosis of the vessel. Injury to or removal of the endothelial cells lining the blood vessels is one of several common denominators inherent to vascular procedures, and current data suggest that spontaneous reendothelialization of these injuries may occur slowly, partially, or not at all.
The endothelial lining of blood vessels is a highly complex, multi-functional cell surface. These cells interact with both the blood and the underlying vessel wall components to maintain a physiological homeostasis. The effects of endothelial injury have been studied in several experimental models mostly designed to study the development of biological mechanisms. After endothelial injury, the vessel wall loses its non-thrombogenic properties. One of the first events to occur is platelet adherence to the vessel surface, which is extensive over the first several days but diminishes over the following week. Steele, P., Chesebro, J., Stanson, A., Holmes, D., Dewanjee, M., Badimon, L., Fuster, V., Circ. Res. 57, No. 1:105-112 (1985). Platelets adhere to the subendothelium and undergo a release reaction, inducing further activation of the plasma coagulation system. Osterud, B. et al., J. Proc. Natl. Acad. Sci. U.S.A., 74, p. 5260 (977). One of the released products is platelet derived growth factor (PDGF), which is mitogenic for vascular smooth muscle cells grown in tissue culture. It has been postulated that local release of this factor may play a role in the genesis of intimal hyperplasia and atherosclerosis. Harker, L. et al., J. Clin. Invest., 58, 731 (1976); Friedman, R. et al., J. Clin. Invest. 60, 1191 (1977). Other substances released from platelets include heparitinase and platelet factor 4. The latter protein has high affinity for heparin and has been shown to penetrate into the vascular media after de-endothelialization. Goldgerg, J. D. et al., J. Science. 209, 611 (1980). Macrophages, which are also a rich source of SMC mitogens, are frequently present in the injured area. Gimbrone, M. A. Jr., In: Jaffe, E. A., Editor, Biology of Endothelial Cells, Martinus Nijhoff Publishers, pp. 97-107 (1984). The final response of the injured arterial wall, independent of whether the injury is chemical, mechanical or biological, is characterized by proliferation of cells in the intima to form a fibro-musculo-elastic plaque. Hoff, H., Thromb. Haemostas., 40, 121 (1970).
Clearly, the endothelial cell plays a key role in the etiology of blood vessel dysfunction. It is anticipated that restoration of intact endothelium immediately following injury might reduce or alter the events occurring immediately after injury. Therefore, it is an object of this invention to provide a method for reendothelializing the linings of vascular passages which have been substantially denuded of endothelial cells.
Recent years have seen refinements made in the isolation of endothelial cells (EC) and their growth in culture. The addition of endothelial cell growth factor (ECFG) and heparin to culture medium has allowed human adult large vessel EC to remain in culture for greater than 50 population doublings. Jaffe, E. A. et al., J. Clin. Invest., 52:2745 (1973); Maciag, T., et al., J. Cell Biol., 91:420 (1981); Thornton, S. C. et al., Science, 222:623-624 (1983); Jarrell, B. E., et al., J. Vasc. Surg., 1:757-764 (1984). Human microvessel EC have also been routinely isolated in large quantities using collagenase digestion and Percoll gradient purification followed by long term cultivation in heparin - ECFG supplemented medium. Jarrell, B. E. et al., Surgery, Vol. 100, No. 2, pp. 392-399 (August 1986).
These advances in EC isolation and culture have been used to better understand the interactions between these EC and prosthetic vascular grafts. Watkins, M. T. et al., J. Surg. Res., 36:588-596 (1984); Williams, S. K. et al., J. Surg. Res., 38:618-629 (1985); Baker, K. S. et al., Am. J. Surg., 150:197-200 (1985). In these studies it has been noted that human EC possess the ability to firmly adhere to both plasma coated surfaces and human amnion type IV/V collagen after a ten to thirty minute incubation period. Jarrell, B. E. et al., Ann. Surg., Vol. 203, No. 6, pp. 671-678 (June 1986). Another observation was that freshly isolated human microvessel EC obtained from fat tissue also possessed this property and could be isolated in quantities of 10.sup.6 EC per gram of fat. Radomski, J., et al., J. Surg. Res., 42, 133-140 (1987); Jarrell, B. E. et al., Surgery, Vol. 100, No. 2, pp. 392- 399 (August 1986).
At least one study has examined the utility of treating the neointimal hyperplasia developed after endarterectomy of a normal artery by endothelial cell sodding. Bush, Jr., Harry L., Jakubowski, Joseph A., Sentissi, Joanna M., Curl, Richard G., Hayes, John A., and Deykin, Daniel, "Neointimal Hyperplasia Occurring After Carotid Endarterectomy in a Canine Model: Effect of Endothelial Cell Seeding vs. Perioperative Aspirin, Journal of Vascular Surgery, Vol. 3, No. 1, pp. 118-125 (January 1987). In this study, endothelial cells were harvested from the veins of dogs selected to undergo treatment. The cells were suspended in sterile autogenous serum, and this suspension was injected into the endarterectomized segment of the dog's artery. From this work, the authors concluded that sodding the endarterectomized surface with autogenous endothelial cells did minimize proliferative lesion in the artery.